Microtubules form specialized arrays with important functions in cell division, cell migration, trafficking and cell morphogenesis. Radial arrays of centrosome-anchored microtubules are typical for proliferating and migrating animal cells. However, differentiated animal cells, as well as all plant cells, can organize their microtubules without centrosomes. Although noncentrosomal microtubule arrays have been implicated in vital cellular functions, including intracellular transport and cell polarity, very little is known aboutthe mechanisms organizing microtubules in the absence of centrosomes. In the course of our preliminary studies we found that microtubule arrays in the model plant Arabidopsis thaliana are regulated by the evolutionally conserved protein phosphatase PP2A complex that includes B regulatory subunit TON2. This proposal will elucidate the PP2A dependent molecular mechanisms of noncentrosomal microtubule array formation. The first aim will establish the functional relationship between the PP2A dependent signaling pathway and the microtubule associate protein TON1A, which was identified in our preliminary studies as a putative TON2/PP2A effector. Functions of TON1A in microtubule nucleation and stability will be determined, and the role of TON2/PP2A signaling in regulation of the TON1A phosphorylation will be addressed. Proposed experiments will provide important insights into mechanisms regulating microtubule nucleation and stability during microtubule array formation. The second aim of the project is to identify and characterize components of the TON1A/TON2/PP2A pathway using a sensitized genetic screen. This will set the stage for a comprehensive understanding of PP2A dependent mechanisms regulating formation of noncentrosomal microtubule arrays.